血管系统的精确建成是胚胎发育的重要事件，形成有效管腔是其中的关键环节，探索其形成机制是血管发育的研究热点。我们前期研究发现斑马鱼CRIP1基因在尾静脉血管丛、肾管等管腔结构中表达，其蛋白与Flk1共存于血管上，敲除后血管发育异常，尾静脉血管丛无法形成正常的静脉, 提示其在血管发育中起重要作用，但作用机制尚不明确。我们拟：首先确定CRIP1的亚细胞定位；在突变体中通过活体成像观察血管发生、发育并检测血管标志性基因表达情况；在哺乳动物细胞中通过细胞转染和RNAi确定CRIP1是否调控细胞迁移及血管生成；筛选与CRIP1相互作用的蛋白，重点关注其中影响血管发育或细胞极性的蛋白，确定CRIP1与候选作用蛋白是否共定位、共注射实验明确在血管发育过程中的功能互作。本研究将从细胞、分子、生化多层次揭示CRIP1在血管发育中的功能及作用机制，构建血管发育调控网络，为血管发育异常疾病的分子机制提供重要参考。

Formation of a cohesive, seamless and contiguous network of blood vessels carrying blood is essential for proper function of cardiovascular system. However the molecular and cellular mechanisms driving the formation of vascular tubes remain to be fully elucidated. The advantage that the zebrafish embryos can survive for about seven days in the absence of blood flow in combination with the embryo transparency has made zebrafish an excellent model to study blood vessel formation. .Very recently, we have characterized a CRIP1 gene in zebrafish, which belongs to group 2 LIM proteins gene family, and encodes a highly conserved protein. Whole-mount in situ hybridization showed that CRIP1 is specifically expressed in posterior cardinal vein plexus as well as pronephric tubule. We also showed that CRIP1 protein is co-localized with the Flk1, a blood vessel marker, in the posterior blood vessels. Moreover, knockout of CRIP1 by Talens leads to abnormal blood vessel formation, in specific, caudal vein plexus failed to consolidate into a single lumen. All these preliminary results suggest that CRIP1 is required for the formation of vascular tubes. However, its precise role and mode of action remain enigmatic, and demand further detailed study. .The aim of this proposal is thus to explore the function and mechanisms of CRIP1 in blood vessel formation. We will first study subcellular localization of CRIP1. Then we will analyze the implication of CRIP1 in the formation of blood vessels by TALENs-mediated gene knockout; the blood vessels phenotype in mutants will be determined by in vivo time-lapse imaging, and the expression of vessel marker genes will be analysed by in situ hybridization. We will also examine whether the vascular system function properly in CRIP1 mutants. Furthermore, we plan to test whether CRIP1 could affect the migration and angiogenesis in vitro in human cell line. To provide a molecular mechanism of CRIP1 action, we project to identify putative CRIP1 protein partners, and to analyze the biochemical and functional interaction between CRIP1 and its interaction partners. All together, these experiments should help to elucidate the mechanism of CRIP1 in the formation and function of blood vessels. They will also provide novel insights into the gene regulatory network underlying blood vessel formation.